Magnetic and Folate Functionalization Enables Rapid Isolation and Enhanced Tumor-Targeting of Cell-Derived Microvesicles

Abstract

Cell-derived microvesicles (MVs), which are biogenic nanosized membrane-bound vesicles that convey bioactive molecules between cells, have recently received attention for use as natural therapeutic platforms. However, the medical applications of MV-based delivery platforms are limited by the lack of effective methods for the efficient isolation of MVs and the convenient tuning of their targeting properties. Herein, we report the development of magnetic and folate (FA)-modified MVs based on a donor cell-assisted membrane modification strategy. MVs inherit the membrane properties of their donor cells, which allows them to be modified with the biotin and FA on their own membrane. By conjugating with streptavidin-modified iron oxide nanoparticles (SA-IONPs), the MVs can be conveniently, efficiently, and rapidly isolated from the supernatant of their donor cells using magnetic activated sorting. Moreover, the conjugated magnetic nanoparticles and FA confer magnetic and ligand targeting activities on the MVs. Then, the MVs were transformed into antitumor delivery platforms by directly loading doxorubicin via electroporation. The modified MVs exhibited significantly enhanced antitumor efficacy both in vitro and in vivo. Taken together, this study provides an efficient and convenient strategy for the simultaneous isolation of cell-derived MVs and transformation into targeted drug delivery nanovectors, thus facilitating the development of natural therapeutic nanoplatforms.

Modularized Extracellular Vesicles: The Dawn of Prospective Personalized and Precision Medicine

“…Based on the fact that 1,2‐dioleoyl‐ sn ‐glycero‐3‐phosphoethanolamine–poly(ethylene glycol) (DSPE–PEG), an amphiphilic molecule, can self‐assemble into lipid bilayers, Zhang et al obtained biotin‐ and folate‐ (FA)‐modified EVs (FA/biotin‐EVs) by culturing the source cells in medium supplemented with biotinylated DSPE–PEG (DSPE–PEG–Biotin) and FA‐modified DSPE–PEG (DSPE–PEG–FA). After conjugation with streptavidin‐conjugated iron oxide nanoparticles, the FA/biotin‐EVs were isolated from culture medium using a magnetic field and demonstrated magnetic steering targeting ability, while magnetic nanoparticles have great potential in diagnosis and treatment 102, 103, 104. We believe that such amphiphilic molecules will be at the frontier of targeted delivery of EVs, as well as studies of isolation and tracing.…”

Preparation of Peptide and Recombinant Tissue Plasminogen Activator Conjugated Poly(Lactic-Co-Glycolic Acid) (PLGA) Magnetic Nanoparticles for Dual Targeted Thrombolytic Therapy

“…When compared with the single targeting strategy, nanoparticles engineered with dual targeting mechanism, either augment the single targeting ligand with a second ligand or with magnetic targeting, has successfully been used for cancer therapy, but it has never have been demonstrated for targeted thrombolysis in vivo. Because dual-targeting nanoparticles were shown to increase differentiation between cancer and normal cells and lead to enhanced accumulation of the drug in tumors [29], the same principle is deemed to be feasible for more selectively targeted delivery of thrombolytic agents to thrombus. Specifically, magnetic PLGA nanoparticles have been explored in a dual targeted delivery of paclitaxel and curcumin in brain tumor therapy [30] and protein antigen delivery for immune stimulation [31].…”

Engineered Extracellular Vesicles as a Reliable Tool in Cancer Nanomedicine

“…Otherwise, the loading of hydrophilic molecules inside the intraluminal space requires the mechanical or chemical disruption of the lipid envelope. Electroporation is based on the application of an electric field to the EVs solution to create nanosized pores in the vesicles' phospholipidic membrane, enabling the diffusion of the desired drug [59,79,86], small interfering DNA (siRNA) [87,88], DNA [89] or NPs [90], maintaining the biological activity of the cargo. However, electroporation can change the physical characteristics of EVs and it is applicable only to small molecules, which can also aggregate and stick on the EVs' surface [85].…”

“…Based on this consideration, if DSPE-PEG is bound to the molecule of interest, it can be incorporated inside the cell membrane, making it overexpresses the molecule on its surface and producing EVs with the desired functionalization. The most frequently used molecules are biotin and folate: the first one binds selectively with streptavidin, used for further functionalization, and the second one targets specific cancer cells [86,[145][146][147]. In addition to folate, also other binding sites can be created on EVs using this approach, for example by adding the RGD sequence or sulfhydryl groups [148].…”

“…Monitoring the uptake by cancer cells [150] Exosomes from macrophage Arginyl-glycyl-aspartic acid (RGD)-functionalized DSPE-PEG (DSPE-PEG-RGD), sulfhydryl-functionalized DSPE-PEG (DSPE-PEG-SH) and folic acid Targeting Hela cells [148] EVs from squamous cell carcinoma DSPE-PEG-Biotin and folate Targeting breast cancer for diagnosis and therapy [145] Exosomes from HUVEC DSPE-PEG-biotin Targeting hepatocellular carcinoma [147] EVs from macrophage DSPE-PEG-Biotin and folate Targeting Hela cells [86] EVs from HUVEC DSPE-PEG-Biotin Targeting melanoma [146] Exosomes from HEK 293T DARPins Targeting HER-2 over-expressing cancer cells (breast, ovarian and gastric cancers) [139] Exosomes from HEK 293 Anti-HER2 scFv antibody (ML39) Inhibit the growth of HER2 positive breast cancer [140] Exosomes from murine melanoma Streptavidin-lactadherin fusion protein linked with biotinylated pH-sensitive fusogenic GALA peptide Cancer immunotherapy [138] Extracellular vesicles from murine neural stem cells Anti-EGFR fused to GPI anchor signal peptides Targeting of Hela cells [143] Exosomes from dendritic cells Lamp2b fused with iRGD (CRGDKGPDC) targeting peptide for αv integrin Targeting breast cancer [79] Exosomes from dendritic cells Carcinoembryonic antigen or HER2 linked to the C1C2 domain of lactadherin Targeting breast cancer [141] Exosomes from HEK 293F cells Prostate-specific antigen, and prostatic acid phosphatase linked to the C1C2 domain of lactadherin Targeting prostate cancer [142] Exosomes from fibrosarcoma cells Chicken egg ovalbumin by fusing it to the C1C2 domain of the lactadherin…”